The role of virus-encoded proteins and viral RNA in phloem-dependent movement and phloem unloading was investigated using temperature-sensitive (ts) tobamoviruses. Phloem-dependent movement of ts-tobamoviruses was not prevented by the non-permissive temperature; only a slight, temporal effect on systemic movement of Tobacco mosaic virus Ni2519 (TMV-Ni2519), which encodes a ts-movement protein (ts-MP) and ts-origin-of-assembly (ts-OAS), was detected. Intact viral coat protein (CP) was not essential in the phloem-unloading step using a modified differential temperature treatment (mDTT), in which the lower inoculated part of a plant was maintained at the permissive temperature and the upper part at the non-permissive temperature. Grafted plants with wild-type rootstocks and MP transgenic scions (MP expressed under transcriptional control of the CaMV 35S promoter) supported phloem unloading of Tomato mosaic virus Ls1 (encoding ts-MP) but not that of TMV-Ni2519 when subjected to mDTT. However, TMV-Ni2519 was still capable of being translocated within the vascular system at the temperature at which the MP was non-functional but could not unload from the vascular tissue. These data suggest that functionality of MP and CP is not essential for phloem-dependent movement and phloem unloading of tobamoviruses, and also indicate that viral RNA participates in the subsequent (cell–cell) movement of tobamoviruses after phloem unloading.